Translational regulation of light-harvesting complex expression during photoacclimation to high-light in Chlamydomonas reinhardtii.

When challenged with excess light, the green alga Chlamydomonas reinhardtii responds, in part, by down-regulating light harvesting capacity at Photosystem II while concomitantly reorganising cellular metabolism to increase sink capacity. We examined the role of translational control during different stages of photoacclimation by analysing polysome profiles of two different light-harvesting complex (LHC) genes encoding a major LHCII component (Lhcbm) and CP29 (Lhcb4) plus iron superoxide dismutase (FeSOD), and through measurement of protein synthesis by in vivo labelling. Within two hours following transfer of low-light (LL) acclimated cultures into high-light (HL), Lhcbm transcripts are off-loaded from polysomes indicating a decline in translational initiation. Lhcbm translational repression at two hours was specific since FeSOD mRNA remained associated with polysomes and in vivo labelling showed an increase in overall protein synthesis. After 4 hours HL exposure, however, a global disassembly of polysomes was detected suggesting a general decline in translational initiation. Interestingly, the decrease in polysomes coincides with maximal FeSOD transcript levels emphasizing that transcript profiles do not always accurately reflect gene expression. Within 8 hours after LL to HL shift, polysomes reform and all transcripts examined are loaded back onto polysomes. Disassembly of polysomes was mimicked by hydrogen peroxide application to non-shifted cultures suggesting that production of hydrogen peroxide due to HL-stress may effect the general decline in polysomes observed during HL acclimation.

Evidence for nucleomorph to host nucleus gene transfer: light-harvesting complex proteins from cryptomonads and chlorarachniophytes.

Cryptomonads and chlorarachniophytes acquired photosynthesis independently by engulfing and retaining eukaryotic algal cells. The nucleus of the engulfed cells (known as a nucleomorph) is much reduced and encodes only a handful of the numerous essential plastid proteins normally encoded by the nucleus of chloroplast-containing organisms. In cryptomonads and chlorarachniophytes these proteins are thought to be encoded by genes in the secondary host nucleus. Genes for these proteins were potentially transferred from the nucleomorph (symbiont nucleus) to the secondary host nucleus; nucleus to nucleus intracellular gene transfers. We isolated complementary DNA clones (cDNAs) for chlorophyll-binding proteins from a cryptomonad and a chlorarachniophyte. In each organism these genes reside in the secondary host nuclei, but phylogenetic evidence, and analysis of the targeting mechanisms, suggest the genes were initially in the respective nucleomorphs (symbiont nuclei). Implications for origins of secondary endosymbiotic algae are discussed.

A phylogenetic assessment of the eukaryotic light-harvesting antenna proteins, with implications for plastid evolution.

The light-harvesting complexes (LHCs) are a superfamily of chlorophyll-binding proteins present in all photosynthetic eukaryotes. The Lhc genes are nuclear-encoded, yet the pigment-protein complexes are localized to the thylakoid membrane and provide a marker to follow the evolutionary paths of plastids with different pigmentation. The LHCs are divided into the chlorophyll a/b-binding proteins of the green algae, euglenoids, and higher plants and the chlorophyll a/c-binding proteins of various algal taxa. This work examines the phylogenetic position of the LHCs from three additional taxa: the rhodophytes, the cryptophytes, and the chlorarachniophytes. Phylogenetic analysis of the LHC sequences provides strong statistical support for the clustering of the rhodophyte and cryptomonad LHC sequences within the chlorophyll a/c-binding protein lineage, which includes the fucoxanthin-chlorophyll proteins (FCP) of the heterokonts and the intrinsic peridinin-chlorophyll proteins (iPCP) of the dinoflagellates. These associations suggest that plastids from the heterokonts, haptophytes, cryptomonads, and the dinoflagellate, Amphidinium, evolved from a red algal-like ancestor. The Chlorarachnion LHC is part of the chlorophyll a/b-binding protein assemblage, consistent with pigmentation, providing further evidence that its plastid evolved from a green algal secondary endosymbiosis. The Chlorarachnion LHC sequences cluster with the green algal LHCs that are predominantly associated with photosystem II (LHCII). This suggests that the green algal endosymbiont that evolved into the Chlorarachnion plastid was acquired following the emergence of distinct LHCI and LHCII complexes.

The fucoxanthin-chlorophyll proteins from a chromophyte alga are part of a large multigene family: structural and evolutionary relationships to other light harvesting antennae.

A fucoxanthin-chlorophyll protein (FCP) cDNA from the raphidophyte Heterosigma carterae encodes a 210-amino acid polypeptide that has similarity to other FCPs and to the chlorophyll a/b-binding proteins (CABs) of terrestrial plants and green algae. The putative transit sequence has characteristics that resemble a signal sequence. The Heterosigma fcp genes are part of a large multigene family which includes members encoding at least two significantly different polypeptides (Fcp1, Fcp2). Comparison of the FCP sequences to the recently determined three-dimensional structure of the pea LHC II complex indicates that many of the key amino acids thought to participate in the binding of chlorophyll and the formation of complex-stabilizing ionic interactions are well conserved. Phylogenetic analyses of sequences of light-harvesting proteins shows that the FCPs of several chromophyte phyla form a natural group separate from the intrinisic peridinin-chlorophyll proteins (iPCPs) of the dinoflagellates: Although the FCP and CAB genes shared a common ancestor, these lineages diverged from each other prior to the separation of the CAB LHC I and LHC II sequences in the green algae and terrestrial plants.

THE CHLOROPHYLL-CAROTENOID PROTEINS OF OXYGENIC PHOTOSYNTHESIS.

The chlorophyll-carotenoid binding proteins responsible for absorption and conversion of light energy in oxygen-evolving photosynthetic organisms belong to two extended families: the Chl a binding core complexes common to cyanobacteria and all chloroplasts, and the nuclear-encoded light-harvesting antenna complexes of eukaryotic photosynthesizers (Chl a/b, Chl a/c, and Chl a proteins). There is a general consensus on polypeptide and pigment composition for higher plant pigment proteins. These are reviewed and compared with pigment proteins of chlorophyte, rhodophyte, and chromophyte algae. Major advances have been the determination of the structures of LHCII (major Chl a/b complex of higher plants), cyanobacterial Photosystem I, and the peridinen-Chl a protein of dinoflagellates to atomic resolution. Better isolation methods, improved transformation procedures, and the availability of molecular structure models are starting to provide insights into the pathways of energy transfer and the macromolecular organization of thylakoid membranes.

Sequence analysis of the plastid rDNA spacer region of the chlorophyll c-containing alga Cryptomonas phi.

A 0.8 kb AvaI/SmaI fragment of the plastid genome of the chlorophyll c-containing alga Cryptomonas phi encompassing the rRNA spacer region and flanking genes has been cloned and sequenced. The spacer region between the 16S and 23S rRNA genes is 275 base pairs long, one of the shortest yet reported, and it contains uninterrupted genes for tRNA(Ile) and tRNA(Ala) separated by only two base pairs. The coding regions for tRNAs and rRNAs have been compared with those from cyanobacteria, land plants and other algae and the possible evolutionary relationships discussed.

The small subunit of ribulose-1,5-bisphosphate carboxylase is plastid-encoded in the chlorophyll c-containing alga Cryptomonas phi.

The gene for the small subunit of ribulose-1,5-bisphosphate carboxylase (Rubisco) is located in the large single-copy region of the plastid genome of the chlorophyll c-containing alga Cryptomonas phi. The coding sequence is 417 base pairs long, encoding a protein of 139 amino acids, considerably longer than most other small subunit proteins. It is found 83 base pairs downstream from the gene for the large subunit and is cotranscribed with it. An 18 base pair perfect inverted repeat is located 8 base pairs beyond the termination codon. Sequence analysis shows the gene to be more closely related to cyanobacterial and cyanelle small-subunit genes than to those of green algae or land plants. This is the first reported sequence of a Rubisco small-subunit gene which is plastid-encoded and it exhibits a number of unique features. The derived amino acid sequence shows extensive similarity to a partial amino acid sequence from a brown alga, indicating that this gene will be of major interest as a probe for the small subunit genes in other algae and for determining possible evolutionary ancestors of algal plastids.